Lymphocyte numbers are regulated both by responses to conventional exogenous antigens and by stimulation by endogenous peptide/MHC complexes. This joint regulation allows individuals to maintain a broad repertoire of specificities, allowing responses against a vast array of foreign entities and, at the same time, providing a pattern of memory based on the immunization history of the individual. The study of the process of lymphocyte dynamics that underlies this regulation requires a multidisciplinary approach, aimed both at the molecular underpinnings of the process through which lymphocytes survive and proliferate and a systemics approach to appreciate the overall mechanisms governing total numbers of lymphocytes of distinct phenotype and distinct specificity. Emphasis has been placed on three aspects of this problem: the dynamics of lymphocyte memory, the mechanisms underlying CD4 T cell depletion in HIV infection and the process of homeostatic proliferation. During the past year, it has been shown that primary responses are highly dependent upon the number of precursor cells that can respond to antigenic challenge. Using both real time PCR and flow cytometric analysis to measure the response when TCR transgenic cells are transferred to intact recipients, it has been shown that responses of CD4 T cells are linear only up to ~5000 precursors in the total populations of lymph node CD4 T cells, with an expansion factor on immunization of ~200. When larger numbers of precursors are present, the factor of expansion falls. Thus, the traditional approach of transferring millions of naive TCR transgenic cells to obtain information about primary immune responses cannot be expected to yield results that reflect physiologic responses. Our analysis of responses made by small numbers of transferred cells provides an approach to assess physiologic responses of naive cells. We are currently examining the impact of memory cells on primary responses and determining how regulatory T cells impact physiologic primary responses. Analysis of the ?steady state? proliferation of memory CD4 T cells has yielded a striking and unanticipated finding. Naive CD4 T cells replicate very slowly; considerably <1% take up BrdU when mice receive a 6 hour ?pulse?. By contrast, among CD4 T cells with a memory phenotype, 3-5% take up BrdU in a 6 hour pulse. Furthermore, the cells that are dividing, appear to replicate multiple times during a relatively short interval. When CD44bright CD4 T cells are labeled with CFSE, transferred to syngeneic mice and examined 7 days later, 12 to 25 % of these cells have divided 7 or more times. This suggests either two populations of cells, one dividing and dying rapidly and one dying and dividing slowly or a process of TCR driven burst-like proliferation, perhaps representing ongoing responses either to exogenous antigens or internal peptides. The analysis of this process will give us important information on the ?global? behavior of CD4 T cells in vivo. We have continued the analysis of the behavior of the factors regulating rapid proliferation of naive CD4 T cells on transfer into lymphopenic recipients. Our results indicate that it is not the number of resident memory cells that determines whether newly transferred cells may proliferate but rather the complexity of the repertoire of the resident memory cells. When naive cells are transferred into mice that have a simple repertoire of memory CD4 T cells, a substantial portion of these cells undergoes rapid, TCR dependent replication. These data imply that the immune system plays a premium on developing a population of memory CD4 T cells with a diverse repertoire and suggests that such cells play an important role, possibly in the initiation or mediation of early protective immune responses.